This project focuses on the regulation and function of the MAPK Kinase Kinase MEKK3. MAPK pathways have been shown to be essential for cell growth, differentiation and apoptosis. It is therefore of interest to gain a better molecular understanding i) of all of the component parts that make up MAPK signaling paths, and ii) of the physiologic targets of individual MAPK pathways in normal and diseased states. We originally cloned MEKK3 and determined that it can activate all four major MAPKs, namely ERK, JNK, p38 and ERK5, albeit to different extents. We also discovered that MEKK3, when overexpressed in an active form, is able to arrest growth, protect from some apoptotic insults and can activate NF-kappaB. In addition, MEKK3 has been proposed to have a direct role in activating the IKK kinase complex, the critical complex that phosphorylates and thus causes degradation of the inhibitors of NF-kappaB. We have cloned an adaptor protein that directly links MEKK3 with the IKK kinase complex and we have determined that this adaptor when overexpressed, negatively regulates TNF-mediated activation of the MAPK JNK/SAPK. We have furthermore discovered that MEKK3 as well as the newly cloned adaptor can interact with AKT, a kinase that has been implicated in many survival pathways and which is able to activate MAPK pathways. These data suggest the existence of a large signaling complex in which both the NF-kappaB-activating IKK core complex as well as MAPK-activating AKT, MEKK3 and its adaptor may reside, likely for the purpose of coordinating the activation of several targets in response to specific signal. To explore the in vivo roles of MEKK3 and its adaptor we have made significant progress in generating mice conditionally deficient in MEKK3 and mice lacking the adaptor.